Abstract Multiple sclerosis (MS), an inflammatory demyelinating disease of the central nervous system (CNS), is the most common cause of non-traumatic neurological disability in young adults in the Western Hemisphere. Disease modifying agents (DMA) that deplete lymphocytes, or block their migration to the CNS, have been shown to reduce the frequency of MS relapses in Phase 3 clinical trials. However, none of those drugs are curative and none are effective in all patients. Granulocyte-macrophage colony-stimulating factor (GM-CSF), a myeloid cell growth factor, has emerged as a promising therapeutic target in MS. GM-CSF producing T cells occur at a relatively high frequency in untreated relapsing remitting MS patients, and have been identified in MS brain tissue. Deficiency or neutralization of GM-CSF confers resistance against experimental autoimmune encephalomyelitis (EAE), an animal model of MS. The mechanism of action of GM-CSF in autoimmune demyelination has yet to be definitively demonstrated. In preliminary studies we found that GM-CSF signaling is necessary for the development of chronic neurological deficits in mice with EAE induced by the adoptive transfer of myelin-specific Th17 cells. GM-CSF blockade or deficiency resulted in a decrease in the numbers and percentages of CNS-infiltrating donor T cells and granulocytes, while CD88 (C5a receptor) expressing myeloid cells were enriched. We found that CNS CD88+ monocytes and myeloid dendritic cells (mDC) are poor antigen presenting cells for myelin-reactive T cells. In animal models of cancer and asthma, CD88+ myeloid cells have immunoregulatory properties. Based on these observations, we propose to investigate two potential mechanisms of action of GM-CSF during the effector phase of EAE. In Aim 1, we will test our hypothesis that GM-CSF deficiency accelerates the conversion of pro-inflammatory iNOS+ myeloid cells in EAE infiltrates to an immunosuppressive CD88+ arginase-1+ phenotype. We will compare cell surface marker expression, antigen presenting capacity, cytokine profiles and immunoregulatory properties of GM-CSF receptor deficient (GM-CSFR-/-) versus WT myeloid cells isolated from the CNS of GM-CSFR-/-/ WT ? WT mixed bone marrow chimeric mice at peak EAE. We will also determine whether treatment of GM-CSFR-/- adoptive transfer recipients with CD88 antagonists exacerbates EAE and promotes chronic disability. In Aim 2 we will investigate the role of GM-CSF in the recruitment of neutrophils, that mediate blood-brain-barrier breakdown, to the CNS. We will determine if forced CNS expression of CXCL1 (a neutrophil attracting chemokine), or treatment with recombinant granulocyte-colony stimulating factor (G-CSF), prevents EAE remission in GM- CSFR-/- hosts. The proposed research could increase our understanding of the immunopathogenesis of EAE and MS, and eventually lead to the development of novel myeloid cell modulating drugs for the treatment of MS patients who do not respond to lymphocyte-targeting DMA.